Since the fungal Mg ion ATPase appears to be an electrogenic proton pump consisting of a single polypeptide chain which can easily be obtained in pure form; it is the best system available at this time to study energy utilization coupled to proton gradients. The purpose of this research plan is to establish the proton pumping characteristics of the yeast plasma membrane Mg ion ATPase and to demonstrate the in vivo function of this enzyme in transport processes. The problem will be approached in the following manner: 1) Proton pumping will be characterized in liposomes reconstituted with the purified enzyme. 2) Protein chemistry procedures will be used to study the structure of the proton pump. 3) Cell physiology studies will be done to show the effect of vanadate, a specific inhibitor of the yeast plasma membrane Mg ion ATPase, on the maintainance of a proton gradient and the transport of small metabolites. 4) Vanadate resistant mutants will be isolated and characterized in an attempt to correlate alterations in the structure of the Mg ion ATPase with changes in in vivo functions of the cell. The data obtained in this study will help us to understand the proton pump activity of the mitochondrial Mg ion ATPase and the mechanism of action of the ion translocating ATPases of the mammalian membrane.